Structural Insight into the Rotational Switching Mechanism of the Bacterial Flagellar Motor

Minamino, Tetsuo; Imada, Katsumi; Kinoshita, Miki; Nakamura, Shuichi; Morimoto, Yusuke V.; Namba, Keiichi

doi:10.1371/journal.pbio.1000616

Cited by 93 publications

(137 citation statements)

References 49 publications

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“…We used total internal reflection fluorescence (TIRF) microscopy to visualize the numbers of molecules of FliM-eYFP (FliM fused to yellow fluorescence protein) in individual motors of tethered cells of E. coli spinning exclusively CW or CCW. The CW cells either contained large amounts of CheY-P or were deleted for cheY and expressed a FliG varient locked in the CW state, which does not affect other motor properties (6,7). CCW cells were simply deleted for cheY.…”

Section: Resultsmentioning

confidence: 99%

“…An independent two-sample t test indicated that this difference was significant (P < 0.001). We did not observe significant correlation between FliM numbers and rotational velocities over a range of speeds typical for tethered cells (1)(2)(3)(4)(5)(6)(7)(8).Because the results of the cryo-EM studies might have been biased, we consider the ratio of the average number of FliM subunits in a given motor to that observed for motors in the CW …”

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confidence: 94%

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Mechanism for adaptive remodeling of the bacterial flagellar switch

Lele

Branch

Nathan

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

128

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The bacterial flagellar motor has been shown in previous work to adapt to changes in the steady-state concentration of the chemotaxis signaling molecule, CheY-P, by changing the FliM content. We show here that the number of FliM molecules in the motor and the fraction of FliM molecules that exchange depend on the direction of flagellar rotation, not on CheY-P binding per se. Our results are consistent with a model in which the structural differences associated with the direction of rotation modulate the strength of FliM binding. When the motor spins counterclockwise, FliM binding strengthens, the fraction of FliM molecules that exchanges decreases, and the ring content increases. The larger number of CheY-P binding sites enhances the motor's sensitivity, i.e., the motor adapts. An interesting unresolved question is how additional copies of FliM might be accommodated.Escherichia coli | motility M otile Escherichia coli swim by rotating helical flagellar filaments with tiny, reversible, ion-driven motors (1). Each motor switches between clockwise (CW) and counterclockwise (CCW) states. Modulation of reversal frequency allows the bacterium to swim toward attractants or away from repellents (chemotaxis). CW rotation is promoted by the binding of an activated cytoplasmic response regulator, CheY-P, to the C-ring component FliM. CheY-P binds to FliM and then to FliN (2), triggering a change in the conformation of FliG, a component at the periphery of the C-ring engaged by the stator element MotA.CheY-P serves to link the input of the chemotaxis network (the receptors) to the output (the flagellar motors). An important feature of the chemotaxis network is precise adaptation to chemical stimuli, which involves resetting of the motor output to the prestimulus output, measured in terms of the probability of CW rotation (CW bias ). Adaptation regulates the chemotactic sensitivity and extends the range of signal detection, enhancing chances of survival. Until recently, adaptation was believed to occur only at the level of chemoreceptors (3). Recent experiments revealed that the motor itself has the ability to adapt to varying CheY-P levels by dynamically increasing the content of FliM, a process that is independent of receptor methylation and demethylation (4). This helps the cell match the motor's operating point to the levels of CheY-P set by the chemotaxis signaling pathway. However, the mechanism by which the motor detects CheY-P levels and remodels the FliM ring is unknown (5). Here, we dissect this mechanism.Results and Discussion Effect of Direction of Rotation on FliM Numbers. We used total internal reflection fluorescence (TIRF) microscopy to visualize the numbers of molecules of FliM-eYFP (FliM fused to yellow fluorescence protein) in individual motors of tethered cells of E. coli spinning exclusively CW or CCW. The CW cells either contained large amounts of CheY-P or were deleted for cheY and expressed a FliG varient locked in the CW state, which does not affect other motor properties (6, 7). CCW cells were simpl...

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Section: Resultsmentioning

confidence: 99%

mentioning

confidence: 94%

Mechanism for adaptive remodeling of the bacterial flagellar switch

Lele

Branch

Nathan

et al. 2012

Proc. Natl. Acad. Sci. U.S.A.

128

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“…The switch complex (which corresponds to the predicted C ring [see below]) also consists of FliM and FliN, which form a pentameric FliM-FliN 4 complex (61,179,242,434,609,632,633). FliM is presumably located between FliG and FliN and contains a binding site for the signaling molecule phospho- CheY, which promotes clockwise rotation of the flagellum (62,375,429,434,508,595). A binding site for phospho-CheY was also identified in FliN, but FliN has a relatively minor role in flagellar switching and rotation (480).…”

Section: Power Supplies-the Cytoplasmic Atpase and The Flagellar Motormentioning

confidence: 99%

Protein Export According to Schedule: Architecture, Assembly, and Regulation of Type III Secretion Systems from Plant- and Animal-Pathogenic Bacteria

Büttner

2012

Microbiol Mol Biol Rev

362

482

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SUMMARY Flagellar and translocation-associated type III secretion (T3S) systems are present in most Gram-negative plant- and animal-pathogenic bacteria and are often essential for bacterial motility or pathogenicity. The architectures of the complex membrane-spanning secretion apparatuses of both systems are similar, but they are associated with different extracellular appendages, including the flagellar hook and filament or the needle/pilus structures of translocation-associated T3S systems. The needle/pilus is connected to a bacterial translocon that is inserted into the host plasma membrane and mediates the transkingdom transport of bacterial effector proteins into eukaryotic cells. During the last 3 to 5 years, significant progress has been made in the characterization of membrane-associated core components and extracellular structures of T3S systems. Furthermore, transcriptional and posttranscriptional regulators that control T3S gene expression and substrate specificity have been described. Given the architecture of the T3S system, it is assumed that extracellular components of the secretion apparatus are secreted prior to effector proteins, suggesting that there is a hierarchy in T3S. The aim of this review is to summarize our current knowledge of T3S system components and associated control proteins from both plant- and animal-pathogenic bacteria.

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“…The FliMFliN ring complex has the rotational symmetry that varies from 32-to 36-fold (25). The crystal structures of FliG (26)(27)(28)(29), FliM (30), and FliN (24) and a crystal structure of the FliG-FliM complex (31) have been solved and possible models for their organization have been proposed (28)(29)(30)(31)(32). However, because the resolution of the MS-C ring structure obtained by electron cryomicroscopy and single particle image analysis is still limited (25), these models still remain ambiguous.…”

mentioning

confidence: 99%

Distinct Roles of Highly Conserved Charged Residues at the MotA-FliG Interface in Bacterial Flagellar Motor Rotation

Morimoto

Nakamura

Hiraoka

et al. 2012

Journal of Bacteriology

100

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Structural Insight into the Rotational Switching Mechanism of the Bacterial Flagellar Motor

Abstract: Structural analysis of a clockwise-biased rotation mutant of the bacterial flagellar rotor protein FliG provides a new model for the arrangement of FliG subunits in the motor, and novel insights into rotation switching.

Cited by 93 publications

References 49 publications

Mechanism for adaptive remodeling of the bacterial flagellar switch

Mechanism for adaptive remodeling of the bacterial flagellar switch

Protein Export According to Schedule: Architecture, Assembly, and Regulation of Type III Secretion Systems from Plant- and Animal-Pathogenic Bacteria

Distinct Roles of Highly Conserved Charged Residues at the MotA-FliG Interface in Bacterial Flagellar Motor Rotation

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